During nervous system development, axons migrate along prescribed pathways in the embryo to reach their appropriate synaptic targets (reviewed in Tessier-Lavigne and Goodman, 1996). One mechanism that contributes to accurate pathfinding is chemorepulsion, the guidance of axons away from non-target regions by diffusible chemorepellent factors secreted by non-target cells. Experiments in which axons are confronted with non-target tissues in tissue culture and are repelled by these tissues at a distance have demonstrated the existence of diffusible chemorepellent activities for numerous axonal classes (Pini, 1993; Fitzgerald et al., 1993; Colamarino and Tessier-Lavigne, 1995; Tamada et al., 1995; Guthrie and Pini, 1995; Shirasaki et al., 1996) as well as for migrating neuronal cells (Hu and Rutishauser, 1996). At the molecular level, two families of guidance cues, the netrin and semaphorin families, have been shown to comprise members that can function as chemorepellents. In Caenorhaditis elegans, the netrin UNC-6 is thought to repel axons that migrate away from the netrin source since these axons are misrouted at a certain frequency in UNC-6 mutants; this presumed repulsion appears to be mediated by the candidate receptors UNC-5 and UNC-40, which are members of the immunoglobulin superfamily (Hedgecock et al., 1990; Leung-Hagesteijn et al, 1992; Hamelin et al., 1993; Wadsworth et al., 1996; Chan et al., 1996). Similarly, in vertebrates netrin-1 can repel subsets of motor axons that migrate away from a source of netrin-1 (Colamarino and Tessier-Lavigne, 1994; Varela-Echavarria et al., 1997), a process which might involve vertebrate homologues of UNC-5 and UNC-40, which have been shown to be netrin-binding proteins (Leonardo et al., 1997; Ackermann et al., 1997; Keino-Masu et al., 1996).
The semaphorins are a large family of structurally diverse secreted and transmembrane proteins characterized by the presence of a conserved .about.500 amino acid semaphorin domain at their amino termini (reviewed in Kolodkin, 1996). The family was first described and implicated in axon guidance through antibody perturbation studies in insects (Kolodkin et al., 1992; Kolodkin et al., 1993). The connection of this family to chemorepulsion was made with the purification of chicken collapsing as a factor that can cause collapse of sensory growth cones when added acutely in cell culture (Luo et al., 1993). Collapsin-1 and its mammalian homologues (Semaphorin III, also known as Semaphorin D) are secreted semaphorins that possess in addition to the semaphorin domain an immunoglobulin domain and a highly basic carboxy-terminal domain (Luo et al., 1993; Kolodkin et al., 1993; Messersmith et al., 1995; Puschel et al., 1995). When presented chronically from a point source, collapsin-1/SemaIII/D (hereafter referred to as SemaIII) can repel sensory and sympathetic axons and has been implicated in patterning sensory axon projections into the ventral spinal cord (Messersmith et al., 1995; Puschel et al., 1995, 1996; Behar et al., 1996; Shepherd et al., 1997). Sema E, which is structurally-related to SemaIII, has also been reported to repel sympathetic axons in culture (cited in Varela-Echavarria and Guthrie, 1997). In Drosophila, the secreted semaphorin SemaIII has been implicated as an inhibitor of axon terminal branch formation (Matthes et al., 1995). However, the mechanisms through which semaphorins produce their repellent or inhibitory actions have not been determined.
To elucidate the mechanisms through which semaphorin proteins produce their repulsive actions on axons, we have sought to identify binding proteins for semphorins on the surfaces of sensory axons. Here we identify two classes of semaphorin receptors, SR1 and SR2, expressed by axons whose function is required for the collapse-inducing and repulsive actions of semaphorins.